Racial and Ethnic Disparities in Phthalate Exposure and Preterm Birth: A Pooled Study of Sixteen U.S. Cohorts.

BACKGROUND: Phthalate exposures are ubiquitous during pregnancy and may contribute to racial and ethnic disparities in preterm birth. OBJECTIVES: We investigated race and ethnicity in the relationship between biomarkers of phthalate exposure and preterm birth by examining: a) how hypothetical reductions in racial and ethnic disparities in phthalate metabolites might reduce the probability of preterm birth; and b) exposure-response models stratified by race and ethnicity. METHODS: We pooled individual-level data on 6,045 pregnancies from 16 U.S. cohorts. We investigated covariate-adjusted differences in nine urinary phthalate metabolite concentrations by race and ethnicity [non-Hispanic White (White, 43%), non-Hispanic Black (Black, 13%), Hispanic/Latina (38%), and Asian/Pacific Islander (3%)]. Using g-computation, we estimated changes in the probability of preterm birth under hypothetical interventions to eliminate disparities in levels of urinary phthalate metabolites by proportionally lowering average concentrations in Black and Hispanic/Latina participants to be approximately equal to the averages in White participants. We also used race and ethnicity-stratified logistic regression to characterize associations between phthalate metabolites and preterm birth. RESULTS: In comparison with concentrations among White participants, adjusted mean phthalate metabolite concentrations were consistently higher among Black and Hispanic/Latina participants by 23%-148% and 4%-94%, respectively. Asian/Pacific Islander participants had metabolite levels that were similar to those of White participants. Hypothetical interventions to reduce disparities in metabolite mixtures were associated with lower probabilities of preterm birth for Black [13% relative reduction; 95% confidence interval (CI): -34%, 8.6%] and Hispanic/Latina (9% relative reduction; 95% CI: -19%, 0.8%) participants. Odds ratios for preterm birth in association with phthalate metabolites demonstrated heterogeneity by race and ethnicity for two individual metabolites (mono-n-butyl and monoisobutyl phthalate), with positive associations that were larger in magnitude observed among Black or Hispanic/Latina participants. CONCLUSIONS: Phthalate metabolite concentrations differed substantially by race and ethnicity. Our results show hypothetical interventions to reduce population-level racial and ethnic disparities in biomarkers of phthalate exposure could potentially reduce the probability of preterm birth. https://doi.org/10.1289/EHP12831.

Cumulative live birth rate in women aged ≤37 years after in vitro fertilization with or without preimplantation genetic testing for aneuploidy: a Society for Assisted Reproductive Technology Clinic Outcome Reporting System retrospective analysis.

Objective: To investigate cumulative live birth rates (CLBRs) in cycles with and without preimplantation genetic testing for aneuploidy (PGT-A) among patients aged <35 and 35-37 years. Design: Retrospective cohort study. Setting: Society for Assisted Reproductive Technology reporting clinics. Patients: A total of 31,900 patients aged ≤ 37 years with initial oocyte retrievals between January 2014 and December 2015 followed through December 2016. Interventions: None. Main outcome measures: The primary outcome was CLBR among patients aged <35 and 35-37 years. The secondary outcomes included multifetal births, miscarriage, preterm birth, perinatal mortality, and the time to pregnancy resulting in a live birth. Adjusted odds ratios (aORs) adjusting for age, body mass index, total 2 pronuclei embryos, embryos transferred, and follow-up timeframe. Results: Among patients aged <35 years, PGT-A was associated with reduced CLBRs (70.6% vs. 71.1%; aOR, 0.82; 95% CI [confidence interval], 0.72-0.93). No association was found between PGT-A and CLBRs among patients aged 35-37 years (66.6% vs. 62.5%; aOR, 0.92; 95% CI, 0.83-1.01). Overall, there was no significant difference in the miscarriage rate (aOR, 0.97; 95% CI, 0.82-1.14). Multifetal birth rates were lower with PGT-A (9.5% vs. 23.1%); however, PGT-A was not an independent predictor of multifetal birth (aOR, 1.11; 95% CI, 0.91-1.36). The average time to pregnancy resulting in a live birth was 2.37 months (SD 3.20) for untested transfers vs. 4.58 months (SD 3.53) for PGT-A transfers. Conclusions: In women aged <35, the CLBR was lower with PGT-A than with the transfer of untested embryos. In women aged 35-37 years, PGT-A did not improve CLBRs.

Associations Between Prenatal Urinary Biomarkers of Phthalate Exposure and Preterm Birth: A Pooled Study of 16 US Cohorts.

Importance: Phthalate exposure is widespread among pregnant women and may be a risk factor for preterm birth. Objective: To investigate the prospective association between urinary biomarkers of phthalates in pregnancy and preterm birth among individuals living in the US. Design, Setting, and Participants: Individual-level data were pooled from 16 preconception and pregnancy studies conducted in the US. Pregnant individuals who delivered between 1983 and 2018 and provided 1 or more urine samples during pregnancy were included. Exposures: Urinary phthalate metabolites were quantified as biomarkers of phthalate exposure. Concentrations of 11 phthalate metabolites were standardized for urine dilution and mean repeated measurements across pregnancy were calculated. Main Outcomes and Measures: Logistic regression models were used to examine the association between each phthalate metabolite with the odds of preterm birth, defined as less than 37 weeks of gestation at delivery (n = 539). Models pooled data using fixed effects and adjusted for maternal age, race and ethnicity, education, and prepregnancy body mass index. The association between the overall mixture of phthalate metabolites and preterm birth was also examined with logistic regression. G-computation, which requires certain assumptions to be considered causal, was used to estimate the association with hypothetical interventions to reduce the mixture concentrations on preterm birth. Results: The final analytic sample included 6045 participants (mean [SD] age, 29.1 [6.1] years). Overall, 802 individuals (13.3%) were Black, 2323 (38.4%) were Hispanic/Latina, 2576 (42.6%) were White, and 328 (5.4%) had other race and ethnicity (including American Indian/Alaskan Native, Native Hawaiian, >1 racial identity, or reported as other). Most phthalate metabolites were detected in more than 96% of participants. Higher odds of preterm birth, ranging from 12% to 16%, were observed in association with an interquartile range increase in urinary concentrations of mono-n-butyl phthalate (odds ratio [OR], 1.12 [95% CI, 0.98-1.27]), mono-isobutyl phthalate (OR, 1.16 [95% CI, 1.00-1.34]), mono(2-ethyl-5-carboxypentyl) phthalate (OR, 1.16 [95% CI, 1.00-1.34]), and mono(3-carboxypropyl) phthalate (OR, 1.14 [95% CI, 1.01-1.29]). Among approximately 90 preterm births per 1000 live births in this study population, hypothetical interventions to reduce the mixture of phthalate metabolite levels by 10%, 30%, and 50% were estimated to prevent 1.8 (95% CI, 0.5-3.1), 5.9 (95% CI, 1.7-9.9), and 11.1 (95% CI, 3.6-18.3) preterm births, respectively. Conclusions and Relevance: Results from this large US study population suggest that phthalate exposure during pregnancy may be a preventable risk factor for preterm delivery.

Occupation and Semen Parameters in a Cohort of Fertile Men.

OBJECTIVE: We examined associations between occupation and semen parameters in demonstrably fertile men in the Study for Future Families. METHODS: Associations of occupation and workplace exposures with semen volume, sperm concentration, motility, and morphology were assessed using generalized linear modeling. RESULTS: Lower sperm concentration and motility were seen in installation, maintenance, and repair occupations. Higher exposure to lead, and to other toxicants, was seen in occupations with lower mean sperm concentrations (prevalence ratio for lead: 4.1; pesticides/insecticides: 1.6; solvents: 1.4). Working with lead for more than 3 months was associated with lower sperm concentration, as was lead exposure outside of work. CONCLUSIONS: We found evidence in demonstrably fertile men for reduced sperm quality with lead, pesticide/herbicide, and solvent exposure. These results may identify occupations where protective measures against male reproductive toxicity might be warranted.

Vanquishing multiple pregnancy in in vitro fertilization in the United States-a 25-year endeavor.

The practice of in vitro fertilization has changed tremendously since the birth of the first in vitro fertilization infant in 1978. With the success of early in vitro fertilization programs in the United States, there was a substantial rise in twin births nationwide. In the mid-1990s, more than 30% of in vitro fertilization cycles resulted in twin or higher-order multifetal pregnancies. Since that time, we not only have witnessed improvements in laboratory and treatment efficacy but also have seen a dramatic impact on pregnancy outcomes, specifically regarding twin pregnancies. Because the field evolved and the risks of multifetal pregnancies became more salient, in 2019, the rate of twin pregnancies had dropped to <7% of cycles. This improvement was largely because of technical advancements and revised professional guidance: culturing embryos longer before transfer, improved freezing technology, embryo preimplantation genetic testing, and revised professional guidance regarding the number of embryos to transfer. These developments have led to single-embryo transfer becoming the standard of care in most scenarios. We used national in vitro fertilization surveillance data of all autologous in vitro fertilization cycles from 1996 to 2019 to illustrate trends in the following improved outcomes: autologous embryo transfer cycles involving blastocyst-stage embryos, vitrified embryos, preimplantation genetic testing cycles, total number of embryos being transferred per cycle, and single-embryo transfer usage over time. Among deliveries from autologous embryo transfers, we highlighted trends in singleton births over time and proportion of deliveries involving twins, triplets, quadruplets, or greater. The notable progress in reducing the rate of multifetal pregnancies with in vitro fertilization was largely attributed to a series of technical and clinical actions, culminating in an 80% reduction in the incidence of multiple births without a loss in overall treatment effectiveness.

Predicting personalized cumulative live birth following in vitro fertilization.

OBJECTIVE: To develop in vitro fertilization (IVF) prediction models to estimate the individualized chance of cumulative live birth at two time points: pretreatment (i.e., before starting the first complete cycle of IVF) and posttreatment (i.e., before starting the second complete cycle of IVF in those couples whose first complete cycle was unsuccessful). DESIGN: Population-based cohort study. SETTING: National data from the Society for Assisted Reproductive Technology (SART) Clinic Outcome Reporting System. PATIENT(S): Based on 88,614 women who commenced IVF treatment using their own eggs and partner's sperm in SART member clinics. INTERVENTION(S): Not applicable. MAIN OUTCOME MEASURE(S): The pretreatment model estimated the cumulative chance of a live birth over a maximum of three complete cycles of IVF, whereas the posttreatment model did so over the second and third complete cycles. One complete cycle included all fresh and frozen embryo transfers resulting from one episode of ovarian stimulation. We considered the first live birth episode, including singletons and multiple births. RESULT(S): Pretreatment predictors included woman's age (35 years vs. 25 years, adjusted odds ratio 0.69, 95% confidence interval 0.66-0.73) and body mass index (35 kg/m2 vs. 25 kg/m2, adjusted odds ratio 0.75, 95% confidence interval 0.72-0.78). The posttreatment model additionally included the number of eggs from the first complete cycle (15 vs. 9 eggs, adjusted odds ratio 1.10, 95% confidence interval 1.03-1.18). According to the pretreatment model, a nulliparous woman aged 34 years with a body mass index of 23.3 kg/m2, male partner infertility, and an antimüllerian hormone level of 3 ng/mL has a 61.7% chance of having a live birth over her first complete cycle of IVF (and a cumulative chance over three complete cycles of 88.8%). If a live birth is not achieved, according to the posttreatment model, her chance of having a live birth over the second complete cycle 1 year later (age 35 years, number of eggs 7) is 42.9%. The C-statistic for all models was between 0.71 and 0.73. CONCLUSION(S): The focus of previous IVF prediction models based on US data has been cumulative live birth excluding cycles involving frozen embryos. These novel prediction models provide clinically relevant estimates that could help clinicians and couples plan IVF treatment at different points in time.

Common practices among consistently high-performing in vitro fertilization programs in the United States: 10-year update.

OBJECTIVE: To evaluate similarities and differences in clinical and laboratory practices among high-performing fertility clinics. DESIGN: Cross-sectional questionnaire study of selected programs. SETTING: Academic and private fertility practices performing in vitro fertilization (IVF). PATIENT(S): Not applicable. INTERVENTION(S): A comprehensive survey was conducted of 13 IVF programs performing at least 100 cycles a year and having high cumulative singleton delivery rates for 2 years. MAIN OUTCOME MEASURE(S): Clinical and laboratory IVF practices. RESULT(S): Although many areas of clinical practice varied among top programs, some commonalities were observed. All programs used a combination of follicle-stimulating hormone and luteinizing hormone for IVF stimulation, intramuscular progesterone in frozen embryo transfer cycles, ultrasound-guided embryo transfers, and a required semen analysis before starting the IVF cycle. Common laboratory practices included vitrification of embryos at the blastocyst stage, air quality control with positive air pressure and high-efficiency particulate air filtration, use of incubator gas filters, working on heated microscope stages, and incubating embryos in a low-oxygen environment, most often in benchtop incubators. CONCLUSION(S): Some areas of consistency in clinical and laboratory practices were noted among high-performing IVF programs that are likely contributing to their success. High-performing programs focused on singleton deliveries. As the field of IVF is rapidly evolving, it is imperative that we share best practices in an effort to improve outcomes from all clinics for the good of our patients.

Dual trigger protocol is an effective in vitro fertilization strategy in both normal and high responders without compromising pregnancy outcomes in fresh cycles.

OBJECTIVE: To study the birth rates of normal vs. high responders after dual trigger of final oocyte maturation with gonadotropin-releasing hormone (GnRH) agonist and human chorionic gonadotropin in fresh in vitro fertilization (IVF) cycles in which ovarian stimulation was achieved by a flexible GnRH antagonist protocol. DESIGN: Retrospective cohort study. SETTING: University hospital. PATIENTS: In women <35 years of age, 290 fresh IVF cycles using the dual trigger protocol with day 5 embryo transfers from January 2013 to July 2018 were included. Cycles excluded were those with preimplantation genetic testing, gestational carriers, donor oocytes, and fertility preservation. INTERVENTIONS: IVF with dual trigger. MAIN OUTCOME MEASURES: Clinical pregnancy rate, live birth rate. RESULTS: Comparing normal responders, defined as <30 oocytes retrieved, and high responders, defined as ≥30 oocytes retrieved, the clinical pregnancy rates (67.0% vs. 69.3%, respectively) and live birth rates (60.5% vs. 60.0%, respectively) were not significantly different. No cases of ovarian hyperstimulation syndrome were reported in either group. CONCLUSIONS: Ovarian stimulation by a flexible GnRH antagonist protocol followed by dual trigger yields comparable outcomes between normal and high responders in fresh IVF cycles.

Infection precautions for severe acute respiratory syndrome coronavirus 2 in assisted reproduction centers: dodging an invisible bullet.

The coronavirus disease 2019 pandemic has resulted in many changes in how we interact in society, requiring that we protect ourselves and others from an invisible, airborne enemy called severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Until a vaccine is developed, and it reaches high levels of distribution, everyone must continue to be diligent to limit the viral spread. The practice of assisted reproduction during this pandemic presents unique challenges in addition to the risks identified in general clinical care. The established good tissue practices employed in laboratories are not designed to protect gametes and embryos from an airborne virus, particularly one that may be shed by an asymptomatic staff member. Armed with theoretical risks but lacking direct evidence, assisted-reproduction teams must examine every aspect of their practice, identify areas at a risk of exposure to SARS-CoV-2, and develop a mitigation plan. Several professional fertility societies have created guidelines for the best practices in patient care during the coronavirus disease 2019 pandemic. As we learn more about SARS-CoV-2, updates have been issued to help adapt infection-control and -prevention protocols. This review discusses what is currently known about SARS-CoV-2 infection risks in assisted reproductive centers and recommends the implementation of specific mitigation strategies.

Factors Impacting Fertility Preservation in Adolescents and Young Adults with Cancer: A Retrospective Study.

Purpose: Fertility preservation before therapy is underutilized for those diagnosed with cancer as an adolescent or young adult (AYA). The purpose of this study was to describe factors impacting utilization of fertility preservation consultations and procedures among AYAs at the University of Iowa Health Care (UIHC). Methods: Patients were identified by the oncology registry at UIHC. Disease site, histology, date of diagnosis, sex, race, ethnicity, insurance, and zip code data were gathered by the registrars. UIHC's electronic medical record was queried for fertility preservation consultation. The Reproductive Endocrinology and Infertility clinical database captured information about patients who underwent fertility preservation. Rural-urban commuting area codes measured rurality. Descriptive statistics and multivariate linear probability models were used to predict the probability of fertility preservation consultation and procedure. Results: From 2008 to 2017, 2932 AYAs were treated for an invasive malignancy at UIHC. Of the 440 (15%) who received a fertility preservation consultation, 156 (5%) underwent a fertility preservation procedure. Multivariate analyses showed that AYAs with public insurance coverage, those diagnosed with central nervous system (CNS) disease or melanoma, and those >30 years old at diagnosis had a significant decrease in the percentage point probability of having a consultation. The percentage point probability of undergoing a procedure was decreased for female patients, those with melanoma or carcinoma, those seen by a pediatric-based provider, and those diagnosed after 25 years of age. Conclusion: This study has important implications for practice and policy, particularly regarding insurance coverage and patient and provider characteristics leading to fertility preservation consultations and procedures for AYAs with cancer.

Effect of body weight on early hormone levels in singleton pregnancies resulting in delivery after in vitro fertilization.

OBJECTIVE: To evaluate which clinical characteristics influence early maternal ß-human chorionic gonadotropin (hCG) and progesterone levels in in vitro fertilization (IVF) pregnancies. DESIGN: Retrospective cohort analysis. SETTING: Academic medical center. PATIENT(S): Women with a live birth after single-blastocyst embryo transfer in either a fresh or frozen cycle between 2004 and 2017, comprising 1,282 pregnancies in 1,057 patients. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): The initial human chorionic gonadotropin concentration (ß-hCG1) measured a mean of 10 days (range: 9-12 days) after embryo transfer, the rate of increase in ß-hCG concentrations, and progesterone concentration, with all analyses controlled for number of days between the embryo transfer and the ß-hCG1 measurement. RESULT(S): The clinical factor that positively influenced the ß-hCG1 level in the fresh cycle was the stimulation type (antagonist cycle higher than long agonist cycle). The clinical factors that negatively influenced both fresh and frozen cycle ß-hCG1 were lower embryo quality and increasing body weight. Increasing weight negatively impacted progesterone levels in both fresh and frozen cycles. A 100 lb (45.4 kg) difference in weight was associated with a 34.8% reduction in ß-hCG1 for both fresh and frozen cycle pregnancies. The rate of increase in ß-hCG was unaffected by body weight. A 100 lb (45.4 kg) difference in weight was associated with a 53.3% and a 32.8% reduction in progesterone in fresh and frozen cycles, respectively. CONCLUSION(S): Increasing body weight is associated with significantly lower ß-hCG and progesterone concentrations in early pregnancy after blastocyst single-embryo transfer in both fresh and frozen cycles. Clinicians should consider this when evaluating these hormone levels for prognostic and diagnostic purposes.

National survey of the Society for Assisted Reproductive Technology membership regarding insurance coverage for assisted reproductive technologies.

OBJECTIVE: To assess the attitudes of Society for Assisted Reproductive Technology (SART) members regarding expanding insurance coverage for patients seeking assisted reproductive technologies (ART) and identify some of the factors that may influence such attitudes. DESIGN: An anonymous online 14-question survey of SART membership; 1,556 surveys were sent through the SART Research Portal from June to December 2017. Questions were incremental in scope, beginning with expanding insurance coverage for ART for vulnerable populations (e.g., fertility preservation for cancer, couples with same recessive gene, fertility preservation for transgender individuals) to extending coverage to include patients who were uninsured for ART. Additional questions assessed attitudes about assuming some fiscal responsibility if mandated insurance were contingent on elective single-embryo transfer (eSET) and lower charges in anticipation of increased number of cases. SETTING: Not applicable. PATIENT(S): Not applicable. INTERVENTION(S): Not applicable. MAIN OUTCOME MEASURE(S): Specific response to 14 survey questions. RESULT(S): The overall response rate was 43.4% (675/1,556). A large majority (>95%) favored insurance for fertility preservation for cancer patients and for avoidance of genetic disorders; 62.3% were supportive of infertility insurance coverage for transgender patients; 78% supported expanding insurance for the broadest segment of the general uninsured population; 76.7% supported expanding insurance contingent on eSET; and 51.3% would consider expanding insurance contingent on lowering charge per cycle in general, but only 23% responded as to what lower charge would be acceptable. Three of four factors were shown by multivariable logistic regression to be predictive of attitudes willing to expand insurance: practice setting (academic > hybrid > private), practicing in a mandated state, and higher annual volume of cases (>500 cycles); these had significant increased adjusted odds ratios ranging from 1.7 to 2.9. A fourth factor, the professional role one had in the practice, was not found to be of significant predictive value. CONCLUSION(S): The great majority of respondents were supportive of expanding insurance for specific segments of vulnerable populations with special needs and for the population who are presently uninsured. Furthermore, the majority of respondents would consider expanding insurance coverage contingent on age-appropriate eSET but have concerns about reduced reimbursement. Those most likely to be willing to expand insurance are those who practice in an academic setting or a mandated state and/or have a high annual volume of cases.

High FSH dosing is associated with reduced live birth rate in fresh but not subsequent frozen embryo transfers.

STUDY QUESTION: Do live birth rates (LBRs) differ between fresh embryo transfer (fresh ET) cycles and their subsequent paired frozen ET (FET) cycles, when comparing cycles based on the total FSH dose used during the fresh cycle? SUMMARY ANSWER: When compared to the paired frozen embryo transfer cycles, the LBR in the fresh cycle of the highest total FSH dose group (>2500 IU) was reduced by 38%. WHAT IS KNOWN ALREADY: There may be a negative association with high gonadotropin doses and LBR after fresh ET. It is unknown whether a similar effect is seen in FET cycles, which are done with increasing frequency. STUDY DESIGN, SIZE, DURATION: In this retrospective observational paired study, we studied IVF cycles between 10 January 2005 and 19 September 2015, for all patients who underwent a fresh, autologous IVF cycle that resulted in at least one fresh ET and at least one FET. PARTICIPANTS/MATERIALS, SETTING, METHODS: The study included 862 women, treated in our academic medical centre, who underwent 935 fresh ET and 1274 FET cycles. Cycles were allocated into three groups based on the total gonadotropin dose they received during their fresh IVF cycle: Group 1 (≤1800 IU FSH), Group 2 (1801-2500 IU), Group 3 (>2500 IU). The primary outcome was LBR after fresh ET and its subsequent paired FET(s), as well as LBR among fresh ETs and FETs as independent samples, based on the total FSH dose used. Implantation rates obtained from fresh and FET cycles were also compared. MAIN RESULTS AND THE ROLE OF CHANCE: The unadjusted fresh LBR was similar between Groups 1 and 2 (46.0% [95% CI: 40.4-51.6] versus 43.8% [38.3-49.4], respectively) but significantly lower in Group 3 (34.4% [29.5-39.8]). The unadjusted frozen transfer LBR was similar among all groups (51.4% [46.7-56.1] versus 46.3% [41.3-51.4] versus 47.5% [42.5-52.4], respectively). When logistic regression analysis with generalized estimating equations was used to control for confounders, the adjusted LBR was found to be similar between the groups both for fresh (odds ratio [OR] = 0.97 [95% CI: 0.61-1.56] Group 2 versus Group 1, OR = 0.69 [0.39-1.21] Group 3 versus Group 1) and FET cycles (OR = 0.87 [0.58-1.31] Group 2 versus Group 1, OR = 0.95 [0.58-1.55] Group 3 versus Group 1). However, for Group 3, the adjusted fresh LBR was 38% lower than its subsequent frozen transfer LBR (OR = 0.62 [0.41-0.93]); this was a statistically significant difference, which was not observed in Group 1 (OR = 0.85 [0.56-1.27]) or Group 2 (OR = 0.95 [0.64-1.41]). LIMITATIONS, REASONS FOR CAUTION: This study is a retrospective cohort, with all of the associated inherent biases. WIDER IMPLICATIONS OF THE FINDINGS: Fresh LBR is negatively impacted by a high dose of total FSH used, as compared to the LBR in subsequent paired FET cycles. Frozen transfer LBR seems unaffected by the total FSH dose used in the fresh cycle, suggesting that the endometrium may be adversely affected, probably indirectly, by high dose gonadotropin use in the fresh IVF cycle only. STUDY FUNDING/COMPETING INTEREST(S): No funding source was used for the completion of this project. There are no conflicts of interest.

Differentially expressed genes in preimplantation human embryos: potential candidate genes for blastocyst formation and implantation.

PURPOSE: The aim of this study was to determine which genes and gene pathways are differentially expressed when comparing human blastocysts with cleavage-stage embryos. METHODS: We individually assessed gene expression in preimplantation human embryos at cleavage (n = 3) and blastocyst (n = 3) stages. Gene expression patterns were then validated in publically available datasets and then independently validated in vitro with additional human embryos using TaqMan gene expression assays. Immunolocalization studies were conducted to identify protein expression in intact blastocyst-stage embryos. RESULTS: Compared to cleavage-stage embryos, blastocyst-stage embryos differentially expressed 51 genes (p < 0.001), with overrepresentation in amoebiasis pathways and pathways in cancer. Of these 51 genes, 21 were found to be independently validated in a separate, publically available dataset, with a substantial agreement with our initial findings (κ = 0.8). In an independent set of cleavage- and blastocyst-stage embryos, we validated that six of eight tested genes were differentially expressed (p < 0.05) by RT-qPCR. Immunofluorescence studies documented the presence of two studied proteins in the trophectoderm of blastocyst-stage embryos. CONCLUSIONS: Differentially expressed genes may be implicated in the invasion and proliferation of the early embryo. Our research highlights specific genes that may be further studied for their role in the implantation process and additionally raises questions about localized gene and/or protein expression in the trophectoderm, which could affect protocols for, and interpretation of, trophectoderm biopsies performed in in vitro fertilization cycles.

Successful treatment algorithm for evaluation of early pregnancy after in vitro fertilization.

OBJECTIVE: To evaluate a prospectively implemented clinical algorithm for early identification of ectopic pregnancy (EP) and heterotopic pregnancy (HP) after assisted reproductive technology (ART). DESIGN: Analysis of prospectively collected data. SETTING: Academic medical center. PATIENT(S): All ART-conceived pregnancies between January 1995 and June 2013. INTERVENTION(S): Early pregnancy monitoring via clinical algorithm with all pregnancies screened using human chorionic gonadotropin (hCG) levels and reported symptoms, with subsequent early ultrasound evaluation if hCG levels were abnormal or if the patient reported pain or vaginal bleeding. MAIN OUTCOME MEASURE(S): Algorithmic efficiency for diagnosis of EP and HP and their subsequent clinical outcomes using a binary forward stepwise logistic regression model built to determine predictors of early pregnancy failure. RESULT(S): Of the 3,904 pregnancies included, the incidence of EP and HP was 0.77% and 0.46%, respectively. The algorithm selected 96.7% and 83.3% of pregnancies diagnosed with EP and HP, respectively, for early ultrasound evaluation, leading to earlier treatment and resolution. Logistic regression revealed that first hCG, second hCG, hCG slope, age, pain, and vaginal bleeding were all independent predictors of early pregnancy failure after ART. CONCLUSION(S): Our clinical algorithm for early pregnancy evaluation after ART is effective for identification and prompt intervention of EP and HP without significant over- or misdiagnosis, and avoids the potential catastrophic morbidity associated with delayed diagnosis.

Lack of carbon air filtration impacts early embryo development.

PURPOSE: To assess human fertilization and preimplantation embryo development in the presence and in the absence of carbon filtration METHODS: This is a retrospective cohort analysis of fresh, controlled ovarian hyperstimulation cycles as well as previously cryopreserved pronuclear stage embryo transfer cycles in a single IVF center. Embryo development and cycle-based outcomes were compared among three groups: 1) when carbon filtration was present, 2) when carbon filtration was absent, and 3) when carbon filtration had been restored. RESULTS: A total of 524 fresh cycles and 156 cryopreserved embryo cycles were analyzed. Fertilization, cleavage, and blastocyst conversion rates for fresh cycles all declined during the period of absent carbon filtration and recovered after the restoration of carbon filtration. Cryopreserved embryos that were thawed and cultured during the period of absent filtration did not have changes in cleavage or blastocyst conversion rates compared to periods where carbon filtration was present. Clinical pregnancy and live birth rates were unchanged among the three time periods. CONCLUSIONS: The absence of carbon filtration in an IVF laboratory air handler is associated with poor fertilization and early embryo development for fresh cycles. Because development of previously frozen pronuclear stage embryos was unaffected, the lack of carbon filtration may preferentially affect embryos in the peri-fertilization period. Carbon filtration is an integral part to a successful human in-vitro fertilization laboratory.

Human embryo cryopreservation-methods, timing, and other considerations for optimizing an embryo cryopreservation program.

The contribution of embryo cryopreservation to the birth rate per in vitro fertilization cycle has escalated from a rare subsidy to a vital tool that is called upon to augment the cycle outcome. Embryology laboratories must identify the embryo stage, quality criteria and methodology that will optimize their ability to preserve each embryo's reproductive potential. This chapter reviews the principles of cryopreservation, outcomes based on embryo stage and cryopreservation method and benchmarks that may be employed by the laboratory to measure the performance of their embryo cryopreservation program.

Alcohol and male reproductive health: a cross-sectional study of 8344 healthy men from Europe and the USA.

STUDY QUESTION: Is there an association between alcohol intake and semen quality and serum reproductive hormones among healthy men from the USA and Europe? SUMMARY ANSWER: Moderate alcohol intake is not adversely associated with semen quality in healthy men, whereas it was associated with higher serum testosterone levels. WHAT IS KNOWN ALREADY: High alcohol intake has been associated with a wide range of diseases. However, few studies have examined the correlation between alcohol and reproductive function and most have been conducted in selected populations of infertile men or have a small sample size and the results have been contradictory. STUDY DESIGN, SIZE, DURATION: A coordinated international cross-sectional study among 8344 healthy men. A total of 1872 fertile men aged 18-45 years (with pregnant partners) from four European cities and four US states, and 6472 young men (most with unknown fertility) aged 18-28 years from the general population in six European countries were recruited. PARTICIPANTS/MATERIALS, SETTING, METHODS: The men were recruited using standardized protocols. A semen analysis was performed and men completed a questionnaire on health and lifestyle, including their intake of beer, wine and liquor during the week prior to their visit. Semen quality (semen volume, sperm concentration, percentage motile and morphologically normal sperm) and serum reproductive hormones (FSH, LH, testosterone, sex hormone-binding globulin, and inhibin B and free testosterone) were examined. MAIN RESULTS AND THE ROLE OF CHANCE: The participation rate for our populations was 20-30%. We found no consistent association between any semen variable and alcohol consumption, which was low/moderate in this group (median weekly intake 8 units), either for total consumption or consumption by type of alcohol. However, we found a linear association between total alcohol consumption and total or free testosterone in both groups of men. Young and fertile men who consumed >20 units of alcohol per week had, respectively, 24.6 pmol/l (95% confidence interval 16.3-32.9) and 19.7 pmol/l (7.1-32.2) higher free testosterone than men with a weekly intake between 1 and 10 units. Alcohol intake was not significantly associated with serum inhibin B, FSH or LH levels in either group of men. The study is the largest of its kind and has sufficient power to detect changes in semen quality and reproductive hormones. LIMITATIONS, REASONS FOR CAUTION: The participation rate was low, but higher than in most previous semen quality studies. In addition, the study was cross-sectional and the men were asked to recall their alcohol intake in the previous week, which was used as a marker of intake up to 3 months before. If consumption in that week differed from the typical weekly intake and the intake 3 months earlier, misclassification of exposure may have occurred. However, the men were unaware of their semen quality when they responded to the questions about alcohol intake. Furthermore, we cannot exclude that our findings are due to unmeasured confounders, including diet, exercise, stress, occupation and risk-taking behavior. WIDER IMPLICATIONS OF THE FINDINGS: Our study suggests that moderate alcohol intake is not adversely associated with semen quality in healthy men, whereas it was associated with higher serum testosterone levels which may be due to a changed metabolism of testosterone in the liver. Healthy men may therefore be advised that occasional moderate alcohol intake may not harm their reproductive health; we cannot address the risk of high alcohol consumption of longer duration or binge drinking on semen quality and male reproductive hormones. STUDY FUNDING/COMPETING INTERESTS: All funding sources were non-profitable and sponsors of this study played no role in the study design, in data collection, analysis, or interpretation, or in the writing of the article. The authors have no conflicts of interest.

Human embryos secrete microRNAs into culture media--a potential biomarker for implantation.

OBJECTIVE: To determine whether human blastocysts secrete microRNA (miRNAs) into culture media and whether these reflect embryonic ploidy status and can predict in vitro fertilization (IVF) outcomes. DESIGN: Experimental study of human embryos and IVF culture media. SETTING: Academic IVF program. PATIENT(S): 91 donated, cryopreserved embryos that developed into 28 tested blastocysts, from 13 couples who had previously completed IVF cycles. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Relative miRNA expression in IVF culture media. RESULT(S): Blastocysts were assessed by chromosomal comparative genomic hybridization analysis, and the culture media from 55 single-embryo transfer cycles was tested for miRNA expression using an array-based quantitative real-time polymerase chain reaction analysis. The expression of the identified miRNA was correlated with pregnancy outcomes. Ten miRNA were identified in the culture media; two were specific to spent media (miR-191 and miR-372), and one was only present in media before the embryos had been cultured (miR-645). MicroRNA-191 was more highly concentrated in media from aneuploid embryos, and miR-191, miR-372, and miR-645 were more highly concentrated in media from failed IVF/non-intracytoplasmic sperm injection cycles. Additionally, miRNA were found to be more highly concentrated in ICSI and day-5 media samples when compared with regularly inseminated and day-4 samples, respectively. CONCLUSION(S): MicroRNA can be detected in IVF culture media. Some of these miRNA are differentially expressed according to the fertilization method, chromosomal status, and pregnancy outcome, which makes them potential biomarkers for predicting IVF success.

Change in oocyte yield in repeated in vitro fertilization cycles: effect of ovarian reserve.

OBJECTIVE: To examine factors that affect variability in oocyte yield between consecutive IVF cycles. DESIGN: Retrospective cohort study. SETTING: University-based fertility clinic. PATIENT(S): A total of 292 women starting two IVF cycles within 12 months from 2005 to 2011. INTERVENTION(S): Variables evaluated included female age, body mass index, parity, infertility diagnosis, antral follicle count (AFC), ovarian stimulation protocol, change in stimulation protocol, total dose of gonadotropin used and change in dose between cycles. Possible associations were tested using a log linear regression model. MAIN OUTCOME MEASURE(S): Change in total and mature oocyte yield between cycles and factors that predict this change. RESULT(S): In cycle 1, total and mature oocyte yield were positively associated with increased AFC and negatively associated with total gonadotropin dose. In cycle 2, a significant increase was seen in both total and mature oocytes. There were no significant independent variables that predicted this change in oocyte yield. When stratified into groups based on ovarian reserve, change in oocyte yield between cycles was significant only in patients with normal ovarian reserve (AFC >10). In this group, the only independent variable associated with an increased oocyte yield was an increase in the total gonadotropin dose. CONCLUSION(S): An increase in oocyte yield between cycles was found in women with normal ovarian reserve and was associated with an increased total gonadotropin dose in the second cycle.